The PML nuclear body (PML-NB) is a common target for disruption by DNA viruses. The adenovirus early protein E4 ORF3 is both necessary and sufficient to rearrange PML-NBs from punctate bodies into nuclear tracks. PML-NB disruption has been correlated with the antagonism of both the innate and intrinsic immune responses. This may reflect the fact that PML, the protein responsible for nucleating PML-NBs, as well as several proteins known to localize to the PML-NB, are products of interferon-stimulated genes. I demonstrate that the E4 ORF3 protein is required for adenovirus DNA replication during the interferon (IFN)-induced antiviral state. When cells are pretreated with either IFN-α or IFN-γ, a mutant virus that does not express E4 ORF3 is severely compromised for replication, suggesting that the functional significance of ORF3 track formation is the inhibition of a PML-NB-mediated antiviral mechanism. Replication of the E4 ORF3 mutant virus is rescued following the introduction of E4 ORF3 from evolutionarily divergent adenoviruses, demonstrating functional conservation for E4 ORF3 inhibition of the IFN-induced antiviral state. Furthermore, E4 ORF3 inhibition of an IFN-induced response is unrelated to the inhibition of adenovirus replication by the Mre11-Rad50-Nbs1 DNA repair complex. I propose that the evolutionarily conserved function of the adenovirus E4 ORF3 protein is the inhibition of an innate response to viral infection mediated by the PML –NB and dependent upon PML and Daxx. Knockdown of either of these proteins is sufficient to rescue replication of a virus deleted for the E4 ORF3 protein during the IFN response. Both the HSV1 ICP0 and HCMV IE1 proteins support DNA replication of adenovirus deleted for the E4 ORF3 protein during the antiviral state, suggesting functional conservation of PML-NB disruption among viruses. I show that this innate response is not based solely on transcriptional repression. While early gene transcription is modestly diminished in the absence of E4 ORF3 protein expression, this reduction does not affect early protein function. I propose that in addition to its abilities to repress gene expression, the PML-NB participates in an alternative innate immune mechanism.